The overall objective of this proposal is to develop mouse models to efficiently test vaccine or gene therapy strategies for the prevention or treatment of HIV infection. The lack of functional lymphocytes in C.B- 17 mice homozygous for the severe combined immunodeficiency (scid) mutation has enabled limited engraftment with human lymphocytes and hematopoietic stem cells. NOD/LtSz-scid/scid mice support heightened levels of human hematopoietic cell reconstitution compared with C.B-17-scid/scid mice. Despite such improved engraftment, the function of transplanted human lymphoid cells as well as the growth and development of human hematopoietic stem cells has been limited by the expression of murine major histocompatibility antigens (MHC), residual murine natural killer (NK) cell activity, and by the lack of human hematopoietic cell growth factors. The specific aims listed below focus on the use of genes disrupted by homologous recombination and the use of human growth factor transgenes to maximize the potential of NOD/LtSz-scid/scid mice as hosts for human adult peripheral blood leukocytes (PBL), hematopoietic stem cells, and fetal hematopoietic tissues. The proposed research will manipulate the microenvironment of the immunodeficient NOD/LtSz-scid/scid host to provide an optimal model for human lymphohematopoietic cell engraftment and function. The specific aims are: l. To create new genetic stocks of NOD/LtSz-scid/scid mice carrying genetically-engineered constructs that reduce MHC complex class I or class II expression, eliminate NK cell activity, and provide human hematopoietic growth factors. II. To determine the phenotypic effects of the disrupted genes and growth factor transgenes on the immunological and hematological systems of NOD/LtSz-scid/scid mice. III. To evaluate the abilities of the new genetic stocks to support engraftment with human lymphohematopoietic cells and infection with HIV. IV. To assess the protective effects of HIV vaccines anti HIV-reactive cloned T cell lines (CTL) following engraftment of the new genetic stocks with human PBL. Genes to be crossed individually onto the NOD/LtSz-scid/scid background include: (a) the beta2 microglobulin (B2m) disrupted allele; (b) the MHC class II (I-A-beta) disrupted allele; (c) the invariant chain (Ii) disrupted allele; (d) the perforin (Pfp) disrupted allele; and (e) three linked human transgenes encoding: stem cell factor, interleukin-3, and granulocyte-macrophage colony-stimulating factor. The disrupted B2m gene will prevent cell surface expression of MHC class I molecules, while the disrupted I-A-beta or Ii alleles will prevent expression of functional MHC class II molecules. The absence of host MHC class I or class II proteins will prevent the development of human T cell xenoreactivity and consequent T cell anergy following engraftment of these immunodeficient mice with human PBL. The absence of functional perforin will eliminate murine NK cell activity, thus enhancing human lymphoid and stem cell engraftment and supporting the generation of immune responses of human origin. The hematopoietic growth factor transgenes encode human cytokines required for the growth and development of human hematopoietic progenitor cells. These small animal models will be tested for human lymphoid and hematopoietic stem cell engraftment and will be utilized for the evaluation of HIV vaccines.